Quantitation of Protein-Bound 3-Nitrotyrosine and 3,4-Dihydroxyphenylalanine by High-Performance Liquid Chromatography with Electrochemical Array Detection

Hensley, Kenneth; Maidt, M.L.; Pye, Quentin N.; Stewart, Charles A.; Wack, M.; Tabatabaie, Tahereh; Floyd, Robert A.

doi:10.1006/abio.1997.2281

Cited by 82 publications

(25 citation statements)

References 27 publications

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“…The nitrated BSA was dialyzed against PBS. The nitration procedure converted about 5% of the tyrosine residues in BSA to nitrotyrosine, as determined by the absorbance at 430 nm (20) and also by the nitrotyrosine and tyrosine content of hydrolyzed samples treated with 6 M HCl and analyzed with HPLC by using electrochemical detection (21,22). BSA was also nitrated by the incubation with 10 mM tetranitromethane for 30 min, which resulted in the nitration of Ϸ10% of the tyrosine residues in BSA.…”

Section: Methodsmentioning

confidence: 99%

An activity in rat tissues that modifies nitrotyrosine-containing proteins

Kamisaki

Wada

Bian

et al. 1998

Proc. Natl. Acad. Sci. U.S.A.

262

178

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Homogenates from rat spleen and lung could modify nitrotyrosine-containing BSA. With incubation, nitrotyrosine-containing BSA lost its epitope to a monoclonal antibody that selectively recognized nitrotyrosine-containing proteins. In the presence of protease inhibitors, the loss of the nitrotyrosine epitope occurred without protein degradation and hydrolysis. This activity was found in supernatant but not particulate fractions of spleen homogenates. The factor was heat labile, was sensitive to trypsin treatment, and was retained after passage through a membrane with a 10-kDa retention. The activity was time-and protein-concentration dependent. The activity increased about 2-fold in spleen extracts with endotoxin (bacterial lipopolysaccharide) treatment of animals, suggesting that the activity is inducible or regulatable. Other nitrotyrosine-containing proteins also served as substrates, while free nitrotyrosine and some endogenous nitrotyrosine-containing proteins in tissue extracts were poor substrates. Although the product and possible cofactors for this reaction have not yet been identified, this activity may be a ''nitrotyrosine denitrase'' that reverses protein nitration and, thus, decreases peroxynitrite toxicity. This activity was not observed in homogenates from rat liver or kidney, suggesting that there may also be some tissue specificity for the apparent denitrase activity.

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Section: Methodsmentioning

confidence: 99%

An activity in rat tissues that modifies nitrotyrosine-containing proteins

Kamisaki

Wada

Bian

et al. 1998

Proc. Natl. Acad. Sci. U.S.A.

262

178

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“…The signals for diCl-Tyr appeared in channels 6 -8, with channel 7 as the dominant channel (where a maximum signal response was observed), channel 6 the leading channel and channel 8 the following one. Peak identity for diCl-Tyr in biological samples was confirmed by comparison of its hydrodynamic voltammogram and the two "ratio accuracy" parameters (the leading channel response to the dominant channel response, and the following channel response to the dominant channel response) to those of the standard (58). To add to further identification, standard diCl-Tyr was also spiked into the biological samples and analyzed subsequently for the proportional increment of the added substance.…”

Section: Methodsmentioning

confidence: 99%

Reactions of Hypochlorous Acid with Tyrosine and Peptidyl-tyrosyl Residues Give Dichlorinated and Aldehydic Products in Addition to 3-Chlorotyrosine

Fu¹,

Wang²,

Davies³

et al. 2000

Journal of Biological Chemistry

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The toxicity of hypochlorous acid (HOCl) generated from activated neutrophils has been associated with several pathological processes such as atherosclerosis. Formation of 3-chlorotyrosine (Cl-Tyr) has been used as a marker for assessing the involvement of HOCl in such processes. In this study, we aimed to investigate the formation of Cl-Tyr from reaction of HOCl with tyrosine (both free and peptide-bound) and the fate of Cl-Tyr under such conditions. Tyrosine, N-acetyltyrosine, bovine serum albumin, and human low density lipoproteins were incubated with a range of reagent hypochlorite concentrations for varying periods in 10 mM phosphate buffer (pH 7.4) at 22°C. The reaction products, and several biological samples, were hydrolyzed (in the case of proteins), isolated, and purified by high pressure liquid chromatography and characterized or quantitated by mass spectrometry and NMR. A significant amount of 3,5-dichlorotyrosine (diCl-Tyr) was obtained from the bovine serum albumin, low density lipoprotein, and some biological samples, in addition to Cl-Tyr, indicating that Cl-Tyr competes effectively for HOCl even when tyrosine is present in great excess. Cl-Tyr and diCl-Tyr were also formed from free tyrosine but then reacted further with HOCl. This finding differs from a claim in the literature that Cl-Tyr was not formed in such a system. The further reaction products of ClTyr and diCl-Tyr with HOCl were elucidated as their corresponding mono-and dichlorinated 4-hydroxyphenylacetaldehydes. These results indicate the importance of assessing other products of HOCl action in addition to Cl-Tyr.

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“…These interesting results show that this analytical system allows the assessment of a correlation between the chromatographic response in terms of peak areas and the redox properties of quinones with different structures. Therefore, the optimized method could be applied in pharmacokinetic studies to evaluate oxidative metabolism and redox biochemical processes, as already reported for processes related to aging [26], immune response [27], and many other pathological processes [28,29].…”

Section: Memoquin Analogs With Modifications In the Quinone Ringmentioning

confidence: 97%

Monolithic stationary phase coupled with coulometric detection: Development of an ion‐pair HPLC method for the analysis of quinone‐bearing compounds

Mancini

Bolognesi

Melchiorre

et al. 2007

J of Separation Science

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Memoquin, a recently discovered new drug candidate for the treatment of Alzheimer's disease (AD), is a compound able to interfere with different key targets of AD neurodegeneration. In the present work, the electroactivity of Memoquin, due to the presence of a quinone ring in the molecule, was exploited for the development of a sensitive and selective HPLC chromatographic method with coulometric detection system. For this purpose, a monolithic silica-based stationary phase (Chromolith C18) was coupled with ESA detector and under high flow rate condition (2 mL/min) gave an efficient coulometric detection without a significant backpressure. The monolithic stationary phase and the electrochemical detection were combined to obtain a very fast (less than 7 min) and sensitive analysis of the compound of interest. For this reason, the method was found suitable to determine Memoquin with very high sensitivity (LOD, 0.005 microg/mL; LOQ, 0.016 microg/mL), better than with UV and fluorescence detections, and selectivity resulting potentially suitable for its analysis in biological samples. Moreover, the HPLC method was employed for the separation of Memoquin from other quinone analogs (endowed with different substituent on the quinone ring and length of the lateral chain) and allowed the comparison of the electrochemical properties of the compounds of the series. In the optimized chromatographic conditions it was possible to separate each quinone and to evaluate the influence of the substituent of the quinone ring on the electrochemical properties of the molecule.

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Quantitation of Protein-Bound 3-Nitrotyrosine and 3,4-Dihydroxyphenylalanine by High-Performance Liquid Chromatography with Electrochemical Array Detection

Cited by 82 publications

References 27 publications

An activity in rat tissues that modifies nitrotyrosine-containing proteins

An activity in rat tissues that modifies nitrotyrosine-containing proteins

Reactions of Hypochlorous Acid with Tyrosine and Peptidyl-tyrosyl Residues Give Dichlorinated and Aldehydic Products in Addition to 3-Chlorotyrosine

Monolithic stationary phase coupled with coulometric detection: Development of an ion‐pair HPLC method for the analysis of quinone‐bearing compounds

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